The present invention relates to a method for increasing the yield or quality of product from a plant by altering the expression of sucrose phosphate synthase. In particular, the present invention provides a transgenic cotton plant that has an increased level of sucrose phosphate synthetase relative to a non-transgenic cotton plant. Methods are also provided for increasing the yield or the quality of cotton fiber and the yield of cotton seed produced from a cotton plant. General methods are provided for regulating the thickness of cell walls, for increasing the yield and quality of other plant fibers, for regulating the ratio of cellulose to other dry weight components of the plant, for increasing seed yield, and for increasing tolerance of photosynthetic efficiency to cool night temperatures.
The control of high-rate cellulose production and its regulation by temperature are critical to agriculture, since all plant growth (and hence the production of all food crops) depends on cellulose synthesis to build cell walls throughout the vegetative and reproductive parts of the plant. The cellulose within the primary walls of all cells of the plant body is also of direct industrial importance as a digestible part of animal forage and for manufacture of thickeners, ethanol, and other cellulose-based or cellulose-derived products. Furthermore, plant parts based on secondary cell walls with high cellulose content are contained in or compose economically important plant products, including cotton fibers, wood, and fibers in forage crops. The agronomic productivity and product quality of wood and cotton, as well as other fiber crops such as hemp and flax, are in large part determined by the biosynthesis of cellulose. Therefore, an understanding of the basic regulatory mechanisms of cellulose synthesis and how it responds to temperature stress allows for beneficial changes in crop plants (improved product yield and quality) through genetic engineering.
Since cotton fiber weight is more than 90% cellulose, cotton is one particular crop where enhancing the flow of carbon to cellulose production can increase yield and quality. This will be an especially beneficial outcome if it is achievable under diverse environmental conditions encountered in cotton production fields, including cool night temperatures that hinder cotton fiber development. For example, it is known that cool night temperatures hinder the seasonal yield and quality of cotton fiber (Gipson, xe2x80x9cTemperature Effects on Growth, Development, and Fiber Properties,xe2x80x9d in Mauney, eds., Cotton Physiology, The Cotton Foundation:Memphis, pp. 47-56) because they hinder the rate of cellulose synthesis (Roberts et al., xe2x80x9cEffects of Cycling Temperatures on Fiber Metabolism in Cultured Cotton Ovules,xe2x80x9d Plant Physiol., 100:979-986 (1992)). The ability to manipulate cotton yield and fiber quality parameters and sustain or improve them under diverse and/or stressful environmental conditions will allow for beneficial changes in crop plants (improved product quality) through genetic engineering.
Cotton fiber yield is the most important determinant of the value of the crop to the producer. Reputable cotton breeders have recently pointed out that cotton production has reached a fiber yield plateau, which bodes ill for the financial success of producers given escalating costs. Potential contributors to this problem include the environmental sensitivity of cotton fiber and seed development, the narrow genetic base of commercial cotton, and the recent introduction of transgenic traits such as herbicide and insect resistance through back-crossing with transformed Gossypium hirsutum cv. Coker 312. Coker 312 (C312) is an old cultivar frequently used for transformation because of its high regeneration capacity. Use of genetic engineering to make cotton crop production more stress resistant, to expand the genetic potential of cultivated cotton, and to improve the yield of transformed cotton with diverse novel traits will bring needed increases in crop yield.
Similarly, seed yield is of value to the cotton producer since seeds are sold for oil production and animal feed. Another minor component, the short fuzz fibers on each seed, provides added economic value to the seed crop. Increased seed and fuzz fiber yield without sacrifice of lint fiber yield or quality would help the producer recover more profit per acre of cotton production. As for cotton seed, increased yield of any seed crop will be of major benefit to agriculture.
Improved cotton fiber quality parameters such as micronaire, maturity ratio, length, length uniformity, bundle strength, and single fiber strength are desired by the textile industry to produce increasingly high quality products and to take full advantage of modern spinning technologies. Fiber quality parameters should also be high enough for the cotton producer to avoid price discounts when he sells his crop to the gin. For example, in a short growing season on the Texas Southern High Plains, producers often suffer price discounts due to low micronaire. Increasingly high fiber quality achieved through breeding has become a required standard in the cotton industry, and market forces may change so producers are more routinely rewarded with price premiums for higher quality cotton. Therefore, stabilizing or increasing fiber quality under diverse environmental conditions through genetic engineering will increase the profitablity of cotton crop production and provide a new spectrum of material properties for exploitation by the processing industries.
Other plant fibers, although often of different tissue origin, share structural features in common with cotton fibers in being elongated cells with cellulose-rich walls. Like cotton fibers, other plant fibers of industrial use are required to have high quality as defined by factors such as cellulose content and wall thickness, diameter, fineness (or coarseness), length, strength, durability, uniformity, elasticity, and elongation. There is an optimum range of such parameters for each particular fiber source and industrial use. Taking examples from wood fibers used after pulping in paper production, longer fiber length and higher single fiber elongation both promote higher paper tear strength. In addition, thick fiber walls promote high pulp yield and production of absorbent paper with high tearing resistance. However, thinner fiber walls promote fiber collapse and better inter-fiber bonding that aids production of high quality writing paper. Therefore, there exists a need to control cell wall thickness and other fiber quality parameters in either negative or positive directions in diverse fibers to improve their yield or quality or expand the range of their industrial utility.
Maximizing crop productivity and utility per acre is a key component of sustainable agriculture. Enhanced production of multiple products from the same crop, such as seed and fiber, would be useful. Similarly, it will be an advantage to maximize the possibility of a successful crop harvest, for example by generating plants with stiffer stems that can better resist lodging in the field without sacrificing the yield of a seed crop.
An increasing level of CO2 in the atmosphere is a concern due to predicted association of rising global temperatures. There exists a need for plants that are better able to immobilize CO2 by conversion of it into useful products, especially products that are typically not burned to regenerate CO2.
Cotton leaves assimilate most carbon into starch during the day, and the starch is converted to sucrose at night for translocation to sinks. As just described, cotton fibers are not well adapted to use this sucrose efficiently for cellulose synthesis during cool nights. Therefore, cool nights reduce cotton photosynthetic efficiency during the following warm day (Warner et al., xe2x80x9cResponse of Carbon Metabolism to Night Temperatures in Cotton,xe2x80x9d Agron. J., 87:1193-1197 (1995)), possibly due to hindered use of carbohydrate at night. The resulting leaf carbohydrate accumulation could signal a down-regulation of photosynthetic genes. The excess starch remaining in the leaf after a cool night could be involved in some negative feedback mechanism reducing photosynthetic rates even after re-warming. There is a need to use genetic engineering to alleviate the cool-night-associated inhibition of photosynthesis during the following warm day.
Sucrose phosphate synthase (xe2x80x9cSPSxe2x80x9d) is a key protein involved in carbon metabolism in plants (See FIG. 1). SPS catalyzes the formation of sucrose phosphate from UDP-glucose and fructose 6-phosphate. In the leaf, SPS is important in controlling the partitioning of reduced carbon between starch and translocatable sucrose (Huber et al., xe2x80x9cRole and Regulation of Sucrose-Phosphate Synthase in Higher Plants,xe2x80x9d Annu. Rev. Plant Physiol. Plant Mol. Biol., 47:431-44 (1996)). In growing sink cells, the data in this invention demonstrate that SPS is involved in directing the flow of carbon to cellulose. Its level of activity can regulate the amount of metabolic flux directed toward cellulose synthesis compared to respiration (See FIG. 2). According to this model, SPS within cellulose-storing sink cells can increase sink strength through an enhanced rate of cellulose synthesis by promoting sucrose synthesis in one or both of two cases: (a) if sucrose transported from the leaves is cleaved to release glucose and fructose before or after entering the sink cells; and/or (b) to reuse the fructose released by the activity of sucrose synthase to channel UDP-glucose and fructose to cellulose synthase. A decreased level of SPS activity can decrease sink strength, by analogous mechanisms, in any case where sink filling is affected by sucrose levels.
In tomato, over-expression of SPS has been shown sometimes to cause a 32% increase in total fruit dry weight. This increase was due not to an increase in individual fruit weight, but to a 50% increase in fruit number (Micallef et al., xe2x80x9cAltered Photosynthesis, Flowering, and Fruiting in Transgenic Tomato Plants That Have an Increased Capacity for Sucrose Synthesis,xe2x80x9d Planta, 196:327-334 (1995)). These tomato plants have also sometimes been shown to have increased fresh fruit weight per fruit and increased fruit soluble solids (sugars) (Laporte et al., xe2x80x9cSucrose-Phosphate Synthase Activity and Yield Analysis of Tomato Plants Transformed with Maize Sucrose-Phosphate Synthase,xe2x80x9d Planta, 203:253-259 (1997)). These reports provide no information about seed yield since tomato seeds weigh little compared to tomato fruits and seeds were not separated from fruits for weighing.
It should be noted that although cotton bolls and tomatoes are both classified botanically as fruits, the nature of the fruits and the relative importance of the seeds they contain is very different. Tomato fruits are essentially sacks of primary cell walls filled with water and soluble glucose, fructose, and sucrose as storage carbohydrates. These sugars crystallize upon drying, contributing to fruit dry weight. Within the fruit, tomato seeds are not a significant sink due to their small size, and they have no economic value except for propagation of tomato. The fruit is the major sink in tomatoes; it constitutes almost all of tomato yield and is the only tomato part with significant economic value.
In contrast, the cotton fruit is relatively dry and thin-walled. The fruit itself does not constitute any substantial sink in cotton or contribute to cotton yield. It protects the seeds only until boll opening, after which it withers. The fruit has no or little economic value (as compost). Cotton seeds with attached fiber represent the two major sinks of substantial economic value in the cotton crop. The cotton fiber is an elongated epidermal cell of the cotton seed coat; it is defined botanically as a trichome. Therefore, the two major sinks in seeds are: (1) the cotyledons of the seed embryo that store oil and protein; and (2) the secondary cell walls of the seed epidermal trichomes (cotton fibers) that store insoluble cellulose. Soluble sugars are not stored in any significant quantity in a mature cotton seed or fruit. Cotton seeds with their attached fiber represent all of the yield in the cotton crop. Therefore, cotton, as well as other fiber producing plants, differ significantly from tomato.
Increased total dry weight of vegetative parts of plants over-expressing SPS has been shown in tomato leaves. In the same study, no change was observed in dry weight of stems and root dry weight decreased (Galtier et al., xe2x80x9cEffects of Elevated Sucrose-Phosphate Synthase Activity on Photosynthesis, Assimilate Partitioning, and Growth in Tomato (Lycopersicon esculentum var UC82B),xe2x80x9d Plant Physiol., 101:535-543 (1993)). Tomato leaves do not contain substantial fiber, being composed mainly of mesophyll cells and conducting vascular tissue. The same plants were shown to sometimes have increased dry weight on a whole-plant basis (Ferrario-Mxc3xa9ry et al., xe2x80x9cManipulation of the Pathways of Sucrose Biosynthesis and Nitrogen Assimilation in Transformed Plants to Improve Photosynthesis and Productivity,xe2x80x9d in Foyer, eds., A Molecular Approach to Primary Metabolism in Higher Plants, Taylor and Francis:New York, pp. 125-153 (1997)) and in above-ground parts including leaves plus stems (Laporte et al., xe2x80x9cSucrose-Phosphate Synthase Activity and Yield Analysis of Tomato Plants Transformed with Maize Sucrose-Phosphate Synthase,xe2x80x9d Planta, 203.253-259 (1997)). In potatoes over-expressing SPS, increased total dry weight of tubers has been shown (Shewmaker, xe2x80x9cModification of Soluble Solids Using Sucrose Phosphate Synthase Encoding Sequences,xe2x80x9d PCT International Publication Number WO 97/15678). Potato tubers do not contain substantial fiber. They are composed mainly of parenchyma cells with primary walls that store abundant starch and lesser amounts of protein. The major yield component of potato tubers is starch. All of these reports lack information on the effect of SPS over-expression on cell wall thickness, cellulose content, and fiber and seed yield of plants. However, the absence of demonstrated increase in stem weight argues against increased fiber content in the tomato plants analyzed.
Increased expression of SPS has been shown to exert other beneficial effects in tomato and Arabidopsis. In both species, leaf starch storage is reduced in preference for synthesis of sucrose. In both species, maximal rates of photosynthesis are enhanced, most significantly in elevated CO2 and saturating light (Galtier et al., xe2x80x9cEffects of Light and Atmospheric Carbon Dioxide Enrichment on Photosynthesis and Carbon Partitioning in the Leaves of Tomato (Lycopersicon esculentum L.) Plant Over-Expressing Sucrose Phosphate Synthase,xe2x80x9d J. Expt. Bot., 46:1335-1344 (1995); Micallef et al., xe2x80x9cAltered Photosynthesis, Flowering, and Fruiting in Transgenic Tomato Plants That Have an Increased Capacity for Sucrose Synthesis,xe2x80x9d Planta, 196:327-334 (1995); and Signora et al., xe2x80x9cOver-Expression of Sucrose Phosphate Synthase in Arabidopsis thaliana Results in Increased Foliar Carbohydrate Accumulation in Plants After Prolonged Growth with CO2 Enrichment,xe2x80x9d J. Expt. Bot., 49:669-680 (1998)). However, these reports provide no information related to effects of cool nights on photosynthesis during the warm day.
Thus, there exists a need for a method to control the level of synthesis of cellulose in fiber producing plants, in particular cotton. There exists a need to be able to control the yield and quality of fibers of commercial value, in particular cotton, under diverse environmental conditions. A general need exists to be able to control the synthesis of cellulose and the thickness of cell walls in plants. A general need exists to promote photosynthetic efficiency in plants growing under cool night temperatures. It is important to be able to increase seed yield in crops as well. The present invention addresses those needs and provides improved plants.
The present invention generally relates to a method of controlling the cellulose synthesis in plants to optimize the level of production and quality of the products derived from the plants.
The invention includes the regulation in the cellulose content, thickness, or yield of any plant cell wall of agricultural or industrial use. Such cell walls include typical thin primary cell walls such as those that are digested in forage and those that exist in useful agricultural residues, for example beet root parenchyma cells remaining after sugar extraction that can be converted into thickening agents. Such cell walls include thick walls such as those of collenchyma and xylem parenchyma that can aid plant rigidity or contribute to yield and digestibility of forage or other agricultural products. Such cell walls also include secondary cell walls such as are commonly found in fiber.
In particular, the present invention provides a transgenic cotton plant that has an increased level of sucrose phosphate synthetase relative to a non-transgenic cotton plant.
The invention also provides a method of increasing the yield of a cotton plant by introducing into the cotton plant a chimeric DNA construct that alters the level of sucrose phosphate synthase activity in an amount sufficient to increase the seed and fiber yield of the cotton plant.
The present invention can also be used to increase the quality of cotton fiber produced from a cotton plant by introducing into a cotton plant a chimeric DNA construct that alters the level of sucrose phosphate synthase activity in an amount sufficient to increase the quality of the cotton fiber produced by the cotton plant.
The invention includes a method of increasing tolerance of photosynthetic efficiency to cool night temperatures by introducing into a plant a chimeric DNA that alters the sucrose phosphate synthase activity in an amount sufficient to increase tolerance of photosynthetic efficiency to cool night temperatures.
In yet another embodiment, the invention provides a method of regulating the ratio of cellulose to other dry weight components of the plant by introducing into a plant a chimeric DNA construct capable of altering sucrose phosphate synthase activity in an amount sufficient to regulate the ratio of cellulose to other dry weight components of the plant.
The invention also provides a method of regulating the thickness of cell walls in a plant by introducing into a plant a chimeric DNA construct that alters sucrose phosphate synthase activity in an amount sufficient to regulate the thickness of cell walls.
In yet another embodiment, the invention provides a method of increasing the harvestable yield of fiber from a fiber containing plant by introducing into a plant a chimeric DNA construct that alters sucrose phosphate synthase activity in an amount sufficient to increase the harvestable yield of fiber from a fiber producing plant.
In yet another embodiment, the invention provides a method of increasing the harvestable yield of seed from a seed producing plant by introducing into a plant a chimeric DNA construct that alters sucrose phosphate synthase activity in an amount sufficient to increase the harvestable yield of seed from a seed producing plant.
In yet another embodiment, the invention provides a method of improving the quality of fiber from a fiber producing plant by introducing into a plant a chimeric DNA construct that alters sucrose phosphate synthase activity in an amount sufficient to regulate fiber quality. Such improvement may be exemplified by changes in length, strength, and weight per unit length.